The immune system protects individuals against disease and infection by viruses, bacteria or other infectious agents. The immune system is able to recognize cells of different individuals, including different allogeneic hosts. Ideally, the immune system functions to eliminate an infectious agent from a mammalian host. Specific immune mechanisms are involved in presenting infection with foreign agents, in resolution of such infections, and in control of cancer cells. Resolution of most virus infections (as well as infection with many other intracellular agents) and elimination of cancer cells is the result of a successful cellular (e.g., T.sub.h 1 helper T cell and cytotoxic T cell) immune response. The cellular immune response results from activation of certain lymphocytes known as T cells.
However, when the immune response is not adequate, the infection can become chronic and persist for many years or even the life-time of the infected host, and can result in life-threatening disease.
Many traditional vaccines expose the immune system to a foreign antigen such as an antigen of an infectious agent to elicit an antigen-specific immune response. The immune response is often predominantly humoral, in which the presence of a foreign antigen elicits the production of antibodies specific for the antigen. For example, an infectious agent antigen may elicit the production of antibodies which neutralize the infectivity of the infectious agent or toxin produced by the agent. Examples include polio and hepatitis A and B, measles, Varicella-zoster, parvovirus and rabies virus antigens. Toxic antigens produced by infectious agents include tetanus toxin, botulinum toxin, diphtheria toxin and pertussis toxin. (See Fundamental Virology, Fields et al. eds., 3rd ed., Lippincott-Raven, New York, 1996; and Microbiology, Davis et al. eds., 4th ed., Lippincott, New York, 1990). This is in contrast to a cellular response, e.g. by activated T cells.
Current methods of treatment of chronic virus infections often provide little clinical benefit because they frequently fail to terminate the infection. Such treatments include administration of small chemical compounds, such as nucleoside analogs, or biologically active proteins, such as interferons. For example, interferons suppress hepatitis B virus (HBV) and hepatitis C virus (HCV) during treatment of chronic infection but virus usually returns to pretreatment levels when treatment is stopped. These treatments inhibit virus replication but do not eliminate virus from cells, or virus infected cells from the infected host, and may result in limited disease improvement only during their administration. Effective vaccination seeks to prevent infection or modify disease resulting from infection caused by the infectious agent to which the vaccine is directed.
Cancer is also a result of failure of the cellular immune system of a patient to eliminate the offending cells, e.g. cancer cells. The cellular immune response plays a major role in the elimination of murine tumors (Wunderlich et al., Principles of Tumor Immunity, In: DeVita et al. (eds.), Biologic Therapy of Cancer, Philadelphia, J.B. Lippincott Co., pp. 3-21, 1991) as it does with infecting viruses. Tumor infiltrating lymphocytes (TILs) that recognize unique cancer antigens in an MHC-restricted fashion have been identified in patients with melanoma. (Rosenberg, J. Clin. Oncol. 10:180-199 (1993)). Most approaches for immunizing patients with cancer have been directed at stimulating strong T cell immune reactions against tumor associated antigens. These studies indicate that adoptive transfer of T lymphocytes from immune animals can transfer resistance to tumor challenge, and in some cases, result in the elimination of established cancer.
A key attribute of the immune system is its ability to discriminate between self and non-self ("foreign"). Optimally, the mammalian immune system is non-reactive ("tolerant") to self-antigens. The mechanisms that provide tolerance eliminate or render inactive clones of B and T cells that would otherwise carry out anti-self reactions. Autoimmune diseases such as multiple sclerosis, rheumatoid arthritis, lupus erythematosus, and Type 2 diabetes mellitus represent an aberrant immune attack in which antibodies or T cells of a host are directed against self-antigen not normally the target of the immune response. Autoimmunity results from the dysfunction of normal mechanisms of self-tolerance that prevent the production of functional self-reactive clones of B and T cells. Attempts to treat such diseases by suppression of the immune response to date have used methods of immunosuppression that are not antigen specific and cause undesirable side effects such as broad suppression of immune responses including those needed for protection against infectious diseases.
In transplant rejection, certain HLA class I protein products ("transplantation antigens") (HLA-A, B and C) of the donor tissue are recognized by antibodies and/or T cells. In allergic responses IgE is produced in response to activation of T.sub.h 2 lymphocytes by antigenic substances or "allergens." Treatments for these reactions have generally involved non-antigen specific suppression of the immune response. More recently new approaches such as use of allergen-specific IgG blocking antibodies for competing with IgE antibodies, and activation of antigen-specific suppressor T cells for inducing anergy in T.sub.h 2 lymphocytes have been investigated.
Methods of antigen specific immune suppression directed at the response to the antigens involved in autoimmune disease ("autoantigens"), transplant rejection and allergic responses are preferred therapeutic approaches to treating these immune disorders.
The mammalian immune system includes B cells and different classes of T cells. CD8+ T (commonly cytotoxic T) cells recognize antigen peptide plus MHC class I complex. Appropriate T cell activation can result in antigen-specific cytotoxic T lymphocytes (CTLs) able to kill target cells expressing the specific viral or cancer antigens.
CD4+ (predominantly helper) T cells recognize antigen peptide plus MHC class II complex. CD4+ T helper cells are classified into two distinct subsets, the T.sub.h 1 and T.sub.h 2 cells, based on their function and pattern of lymphokine secretion. T.sub.h 1 cells secrete principally IL-2, IL-12, IFN.gamma. and TNF (providing help for the activation of CTLs and an anti-viral or anti-tumor response), while T.sub.h 2 cells principally secrete IL-4, IL-S, IL-6 and IL-10 (providing help for the humoral immune response, such as antigen specific B cell proliferation, differentiation and maturation). The T.sub.h 1 type cytokines enhance cellular immunity and have anti-viral or anti-tumor activity. The T.sub.h 2 type cytokines enhance antibody production, particularly IgE production leading to stimulation of allergic responses, and help respond to extracellular infections.
The differentiation of the CD4+ cells into T.sub.h 1 or T.sub.h 2 subsets during the activation process can be influenced not only by the antigenic stimulus but also by the presence of cytokines. Thus, the T.sub.h 1 or T.sub.h 2 response can be influenced by exposure to molecules such as cytokines that favor one type of response over the other. Factors that are known to enhance the T.sub.h 1 response include intracellular pathogens, exposure to IFN, IL-12 and IL-2, the presence of professional APCs and sustained exposure to low doses of antigen. Factors that are known to enhance the T.sub.h 2 response include exposure to IL-4 and IL-10, APC activity on the part of B lymphocytes and high doses of antigen.
T lymphocytes ("T cells") of the mammalian immune system recognize antigen in the form of short peptides derived ("processed") from the native protein antigen complexed with self glycoproteins encoded by the major histocompatibility complex ("MHC") molecules and transported to the surface of an antigen presenting cell (APC) (Whitton et al. in Virology, 2nd Ed., (Fields et al., eds.), Ch. 15, pp. 369-381, Raven Press, Ltd., New York, 1990). MHC recognition of foreign peptides provide antigen specificity for immune responses. The MHC is a cluster of closely linked genetic loci encoding three different classes of polypeptide products (class I, II and III) involved in the generation and regulation of immune responses. Genes encoding MHC polypeptides are present in all vertebrates. Class I and II MHC genes encode cell-surface proteins involved in the presentation of protein antigens to T cells during generation of an immune response. Antigenic peptide fragments of foreign protein antigens complexed with Class I molecules on the surface of antigen presenting cells can be recognized by T cells. Antigen peptide complexed with MHC on the cell surface renders that cell a target for antigen specific cytotoxic T cells. Some tumor associated protein antigens also bind to Class I molecules on the surfaces of neoplastically transformed cells rendering them targets for antigen specific CTLs. In transplant rejection, foreign class I alleles are recognized by the cytotoxic T cell receptor leading to alloreactivity and lysis.
Class II MHC molecules are primarily expressed on the surfaces of B lymphocytes and antigen-presenting macrophages and on activated T cells and are involved in presenting antigen to helper T cells for generating an immune response. In particular, Class II molecules regulate the activation of antigen-specific MHC-restricted helper T cells required for activation of cytotoxic T lymphocytes and antibody-producing B cells. Normally, class II MHC expression is limited to professional antigen presenting cells such as B lymphocytes, macrophages, dendritic cells and activated T cells in humans that process antigens for T cell activation. In humans, there are at least three types of Class II molecules, HLA-DR, DQ and DP.
The human MHC located on chromosome 6 is referred to as HLA (Human Leukocyte Antigen). The combination of molecular alleles at the clustered HLA loci defines the tissue type ("HLA specificity") of an individual. There are vast polymorphisms in each HLA gene locus and each individual has a personal set of Class I and Class II HLA molecules ("self" HLA). HLA molecules serve an important role in human immune system recognition of "self" from "non-self" or foreign. When cells from a donor are introduced into hosts with different HLA genotypes, the donor cells are recognized as foreign for the donor cell HLA molecules and the donor cells are destroyed by the host immune system.
Another important feature of the antigen specific T cell activation is its MHC restriction. T cells from a donor can only be activated by an antigen in complex with "self" MHC (HLA in human) molecules and the activated CTLs can only kill target cells presenting the same antigen in complex with "self" MHC molecules. Activation of T lymphocytes ("T cells") occurs when the T cell receptor (TCR) binds to an antigen peptide complexed with a self HLA molecule on the surface of APCs.
Activation requires not only recognition of the antigen peptide-MHC complex by the TCR (first signal), but also the interaction of costimulation molecules on the surface of APCs with specific molecules on the surface of T cells (second signal) (Freeman et al., J. Exp. Med. 174:625-691 (1991)). Such costimulation molecules include B7- 1, B7-2 and B7-3 proteins as well as CD40 (Clark, et. al., Ann. Rev. Immunol. 9 97-127 (1991)). In contrast, antigen presentation to T cells in the absence of a second signal, such as the B7-CD28 signal, leads to T cell anergy (tolerance) to the antigen. (Boussiotis et al., Immunol. Revs. 153:5-26 (1996); Judge et al., Immunologic Res. 15(1):38-49 (1996); McIntosh et al., Cellular Immunol. 166(1):103-112 (1995); Boussiotis et al., Current Op. in Immunol. 6(5):797-807 (1994); and Gimmi et al., Proc. Natl. Acad. Sci. USA 90(14):6586-6590 (1993)). Antigen-specific tolerance provides a preferred immunosuppressive approach for treatment of auto-immune disease, transplant rejection and allergic reactions.
The level of MHC molecule expression on the cell surface is also an important factor for both the activation and the effector function of T cells. The level of expression of HLA molecules on the cell surface is thought to be regulated by a number of factors. IFN, a lymphokine, can up-regulate the cell surface expression of HLA molecules, enhancing the function of APCs to activate T cells (Revel, et. al., Trends Biochem Sci 11 166 (1986)). Other molecules, such as PA28 (Groettrup, et. al., Nature, 381:166-8 (1996)), can up-regulate the surface expression of HLA molecules and the loading of antigenic peptides to the HLA molecules.
Two pathways are thought to exist within vertebrate cells to generate peptides for recognition by T cells. One is the endogenous pathway, which processes endogenously expressed antigenic proteins and provides peptides to MHC class I molecules for antigen presentation to CD8+ T cells. This process involves proteasomes and the ubiquitin pathway of protein degradation. Additionally, specific peptide transporter proteins (TAP) transport the peptides across the membrane of the endoplasmic reticulum (ER) to access the lumen, where antigenic peptide is bound to the class I molecule. A large family of related transporter proteins is known as the "ABC (ATP binding cassette) transporters". TAP transporter is a member of this family. The TAP molecule is composed of two polypeptide chains, TAP1 and TAP2, both of which are encoded by genes in the HLA family. The other is the exogenous pathway, which processes exogenous antigenic proteins and provides peptides to HLA Class II molecules for presentation to CD4+ T cells. Peptide loading to HLA class II molecules requires the presence of a molecule, HLA-DM in humans. The class II locus of the HLA genes contains genes encoding proteins involved in antigen processing including LMP2 and LMP7 (genes for two proteasome subunits) and TAP1 and TAP2, peptide transporter heterodimers, and the HLA-DM molecule (DMA and DMB) (Monaco, J. Leukoc. Biol. 57:543-547 (1995); Howard, Curr. Opin. Immunol. 7:69-76 (1995)). Sufficient cellular levels of expression of such molecules as the peptide transporters and proteasome components are required for cells to present antigen to T cells. The level of expression of HLA molecules is also important for this function.
In recent years, APCs have been contemplated for use as immunogen for cellular therapeutics. There have been a variety of suggestions for modifying cells to provide immunotherapeutics (for review see Tykocinski et al., Amer. J. Pathol. 148:1-16 (1996)). Tumor cells have been designed for use in cellular cancer vaccines, for example by introducing genes encoding proteins with known immunostimulatory properties such as cytokines, chemokines, heat shock proteins and MHC molecules into the tumor cells ex vivo (Ostrand-Rosenberg, Curr. Opin. Immunol. 6:722-727 (1994); Pardoll, Curr. Opin. Immunol. 4:619-623 (1992), Townsend et al., Science 259:368-370 (1993); and Townsend and Allison, Cancer Res. 54:6477-6483 (1994)). However, gene transfer into tumor cells has limitations because most primary tumor cells grow poorly in cell culture and are poor transfection targets.
Several different cell types can function as professional APCs ("PAPCs"), including macrophages, monocytes, dendritic cells, Langerhans cells and activated B cells. These cells express molecules such as costimulatory molecules capable of providing the second signal for T cell activation. Other cells types (non-professional antigen presenting cells), to varying extents, possess antigen processing and antigen presenting capabilities.
While APCs have been obtained from human hosts for manipulation ex vivo and reintroduction into the host ("autologous APCs"), there remains a need for antigen presenting cells that can be used therapeutically to more strongly stimulate antigen specific immune responses or produce antigen-specific suppression of immune responses. Viral antigens can be degraded in infected cells and specific viral antigen peptide fragments presented at the cell surface complexed with MHC molecules where they serve as targets for MHC-restricted CTLs. However, most virus infected cells in the host can not serve as APCs for the activation of T cells because they do not express molecules that can provide a costimulation signal required for T cell activation. When antigen specific T cell activation is inadequate and fails to eliminate the agent, infections may persist and become chronic. Similarly, most cancer cells can serve only as target cells for CTLs, not as professional APCs for activation of T cells. Cancer results when the cancer-specific T cell activation is not sufficient and CTLs fail to adequately kill cancer cells.
There is thus a need for engineered cells having a compatible HLA specificity to that of a subject, so that the cells can be introduced into the subject to present selected antigens and direct the subject's immune response to these antigens for treatment of infectious diseases and cancer, and for antigen-specific immunosupression of unwanted immune responses.